A Comparative Study of Functional Properties of Calf Chymosin and Its Recombinant Forms

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V. V. Starovoitova¹, T. I. Velichko¹, L. A. Baratova², I. Yu. Filippova^1*, and G. I. Lavrenova¹

¹Chemical Faculty, Lomonosov Moscow State University, 119992 Moscow, Russia; fax: (7-495) 939-8846; E-mail: irfilipp@genebee.msu.su

²Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, 119992 Moscow, Russia; fax: (7-495) 939-3181; E-mail: baratova@belozersky.msu.ru

^* To whom correspondence should be addressed.

Received April 5, 2005

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The action of calf chymosin obtained from transgenic sheep milk and the recombinant protein expressed in yeast Kluyveromyces lactis (Maxiren) on fluorogenic peptide substrates, namely Abz-A-A-F-F-A-A-Ded, Abz-A-A-F-F-A-A-pNA, Abz-A-F-F-A-A-Ded, Abz-A-A-F-F-A-Ded, Abz-A-A-F-F-Ded, Abz-A-A-F-F-pNA, and heptapeptide L-S-F-M-A-I-P-NH₂, a fragment of kappa-casein (the native chymosin substrate), was investigated. It has been established that transgenic chymosin and recombinant chymosin (Maxiren) differ from the native enzyme in their action on low molecular weight substrates, whereas there was no difference in enzymatic action on protein substrates. Pepstatin, a specific inhibitor of aspartic proteinases, inhibits the recombinant chymosin forms less efficiently than the native enzyme. Perhaps this is associated with local conformational changes in the substrate binding site of recombinant chymosin occurring during the formation of the protein globule.

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Key words: chymosin, recombinant chymosin, fluorogenic substrates, pepstatin

DOI: 10.1134/S0006297906030138

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